Mobile phase strength, effect

Obtaining data on the effects of these parameters may allow one to judge whether a method needs to be revalidated when one or more parameters are changed. For example, if column performance changes over time, adjusting the mobile-phase strength to compensate for changes in the column may be allowed if such data are included in the validation. 

The concentration of buffer is also a very important aspect in the optimization of the chiral resolution on these CSPs. It has been reported that an increase in buffer concentration caused a decrease in the retention and selectivity for all amino acids except for the basic amino acids. Therefore, the separation of basic amino acids is possible only with the most concentrated buffers. The buffers of concentrations in the 25-50-mM range were used for the chiral resolutions with some exceptions. In spite of this, few reports are available for the optimization of the chiral resolution by varying the ionic strength of the mobile phase. The effect of ionic strength of phosphate buffer on the chiral resolution of serine was carried out by Gubitz and Jellen [18] and the best resolution was achieved at 0.01 M concentration (Fig. 7). In another study, the concentration of ammonium acetate (0.001-0.01 M) was varied to optimize the chiral resolution of amino acids [19]. The effect of the concentration of ammonium acetate on the chiral resolution of amino... 

Another injection-related effect that can diminish the separation performance is the diluent effect, also known as solvent mismatch. This occurs when the elution strength of the sample solvent is greater than the starting mobile-phase strength. The retention of the analyte on the stationary phase is less in the small plug of sample solvent than it is in the surrounding bulk... 

Figure 2.12. Four RPLC chromatograms illustrating the effect of mobile phase strength and selectivity of acetonitrile (ACN) and methanol (MeOH). See Figure 2.10 for LC conditions.

Contamination can be the result of a buildup of retained sample material on the stationary phase. This can be a problem with gradient elution systems when the proportion of organic modifier in the mobile phase increases over the course of the chromatographic run. These unwanted materials can be washed off the column with the increase in mobile phase strength, which in turn can distort the baseline in the form of waves. In some circumstances, this can interfere with late eluting peaks of interest and can have a detrimental effect on the assay outcomes. 

Mobile-phase selection for cationic polymers is similar to that for the other polymers in that ionic strength and pH can change the shape of the solute from linear to globular (9). Mobile phases are often low pH e.g., 0.1% trifluo-roacetic acid, including 0.2 M sodium chloride, has been used successfully for polyvinylpyridines. Sodium nitrate can be substituted for the chloride to avoid corrosive effects. Some salt must be included so that ion exclusion does not occur (3). 

As we continue lowering the pressure, GC is the final limiting case when the mobile phase has zero solvent strength over the entire column length and where temperature is the only effective control parameter. Gas chromatography is shown in Figure 7.3. 

Naphthalenedisulfonate-acetonitrile as the only mobile phase with a silica column coated with a crosslinked aminofluorocarbon polymer has proven to be an effective combination for the separation of aliphatic anionic surfactants. Indirect conductivity and photometric detection modes are used to monitor these analytes. The retention of these surfactants is found to depend on both the ionic strength and the organic solvent content of the mobile phase. The mechanism of retention is considered to be a combination of both reverse phase and ion exchange processes. Selective separation of both alkanesulfonates and... 

Intrinsic viscosity measurements were done with a large number of solvents varying in pH, ionic strength, etc., using Cannon-Ubbelohde semimicro dilution viscometers. This was done to provide information on the effect of mobile phase composition on the size of a polymer molecule in solution and thus to facilitate the interpretation of GPC behavior. 

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